Heme Oxygenase-mediated Resistance to Oxygen Toxicity in Hamster Fibroblasts

Abstract

The role of heme oxygenase (HO)-1 was evaluated in the oxygen-resistant hamster fibroblast cell line, O 2 R95, which moderately overexpress HO when compared with the parental cell line, HA-1. To suppress HO-1 expression, O 2 R95 were transfected with HO-1 antisense oligonucleotide or treated with tin-mesoporphyrin (SnMP). To increase HO-1 expression, cells were transfected with HO-1 cDNA in a pRC/cytomegalovirus (CMV) vector. All cells were challenged with a 48-h exposure to 95% O 2 (hyperoxia). When HO activity was suppressed, O 2 R95 cells had significantly decreased cell viability, increased susceptibility to lipid peroxidation, and increased protein oxidation in hyperoxia. In contrast, further overexpression of HO-1 did not improve resistance to oxygen toxicity. Antisense-transfected cells and SnMP-treated cells with lowered HO activity showed increased levels of cellular heme compared with controls. In the HO-1 cDNA-transfected O 2 R95 cells, cellular heme was lowered compared with controls; however, cellular redox active iron levels were increased. We conclude that HO mediates cytoprotection to oxygen toxicity within a narrow range of expression. We speculate that this protective effect may be mediated in part through increased metabolism of the pro-oxidant heme but that higher levels of HO activity obviate protection by increased redox active iron release.Heme oxygenase (HO-1), 1 the rate-limiting enzyme in the conversion of heme to bilirubin, is known to be induced by various oxidant stresses. However, it is not clear whether HO serves in protection against hyperoxia and, if so, by which mechanisms. HO-1 antisense transfection experiments have shown that higher HO-1 protein levels were associated with protection against UVA radiation (1). Furthermore, transfection of coronary vessel endothelial cells with an overexpression vector containing HO-1 cDNA resulted in resistance against hemoglobin-induced injury (2). We have previously shown that HA-1 hamster fibroblasts made stably resistant to oxygen toxicity (O 2 R95) had 1.8-fold higher HO activity (3), suggesting that moderate increases in HO activity may be beneficial in resistance to oxygen toxicity. However, O 2 R95 cells have increases in other antioxidants that may also contribute to their resistance to oxygen toxicity (4), and no direct evidence currently exists linking HO to resistance to oxygen toxicity.Investigators have hypothesized that HO may serve a role in protection against oxidative injury by forming the antioxidant molecules biliverdin and bilirubin (5, 6). Additionally, induction of ferritin with enhanced HO activity has been observed (1). This could lead to sequestration of redox active iron, thereby conferring protection against oxidative stress (7). Another possible antioxidant mechanism of HO could involve the destruction of heme itself. Heme and hemoproteins have been shown by several investigators to be instrumental in exacerbating oxidative injury (8, 9). This has lead to the hypothesis that reduction of the cellular heme pool by ...

Adenovirus-mediated transfer of heme oxygenase-1 cDNA attenuates severe lung injury induced by the influenza virus in mice

“…15,31 Based on the knowledge that overexpression of HO-1 prevents cell death from a variety of stresses including hyperoxia, exposure to heme, hydrogen peroxide or LPS, we performed TUNEL analysis to detect DNA fragmentation in lung cells as a measure of cell damage. 31,42,43 Consistent with previous findings that influenza virus inoculation results in severe apoptosis of respiratory epithelium, inoculation with H1N1 in our study resulted in a marked increase in the number of respiratory epithelial cells and inflammatory cells with damaged DNA. [44][45][46] In contrast, animals receiving pretreatment of Ad.HO-1 demonstrated marked reduction in cells positive for DNA damage, thus preventing injury.…”

Antiapoptotic effect of haem oxygenase-1 induced by nitric oxide in experimental solid tumour

“…Although several reports have shown a cytoprotective effect of HO-1 (Dennery et al, 1997;, contradictory effects of the biological consequences of overexpression of HO-1 have also been reported. Specifically, a study by indicated that overexpression of HO-1 could exacerbate the oxidative stress of cells, and they revealed a critical role of reactive iron (ferric iron, Fe 2+ ) released during HO-catalytic decomposition of the porphyrin ring of haem in determining the consequence of HO expression.…”

“…Haem oxygenase -1 (HO-1), the inducible isoform of HO, also known as heat shock protein 32 (HSP32), is found at low levels in most mammalian tissues, but it is constitutively expressed in liver and spleen and is upregulated by its substrate haem (Tenhunen et al, 1970) and various stress-inducing stimuli such as UV light (Keyse and Tyrrell, 1989), heavy metals (Mitani et al, 1993), heat shock (Shibahara et al, 1987), hypoxia (Motterlini et al, 2000), and nitric oxide (NO) (Kim et al, 1995;Foresti and Motterlini, 1999;Bouton and Demple, 2000). Induction of HO-1 is suggested to have a cytoprotective effect against oxidative injury (Dennery et al, 1997;, because some bile pigments formed via HO have been shown to behave as antioxidants (Minetti et al, 1998;Dore et al, 1999).…”

Overexpression of heme oxygenase-1 protects smooth muscle cells against oxidative injury and inhibits cell proliferation

“…As previous report, HO-1 is proposed as a protective protein as heat shock protein that is inducible by a number of stressful stimuli, such as hypoxia, heavy metals, oxidant-LDL and endotoxin. It was suggested in the considerable data that the increase in HO-1 expression in different cell types such as endothelial cells [23], cardiomyocytes [24], fibroblasts [25] etc may yield protective effect on the insulated cells against oxidation injury. Clark [26] reported that hemin-mediated increase in HO-1 protein expression resulted in high resistance to VSMC injury caused by an oxidant-generating system, but expression of HO-1 induced by hemin pre-treatment appeared non-tissue specific.…”